This application is in response to RFA-AI-05-042 on Innate Immunity to Category B protozoa of which Cryptosporidium spp rank among the most significant. Two species, C. hominis and C. parvum, are linked with human cryptosporidiosis, a serious cause of morbidity and mortality worldwide, and against which there is no effective therapy of prevention. Our goal is to elucidate the mechanisms by which immune cells initiate resistance against cryptosporidiosis with a view that a better understanding of the various components of the immune response will lead to development of effective vaccines and adjuvants to combat the infection in humans. Our central hypothesis is that dendritic cells recognize Cryptosporidium through Toll like receptor(s) which initiate the process of resistance against parasite invasion. We base this on the observations that: 1) dendritic cells sense pathogens through Toll-like receptors which lead to the initiation of adoptive immune responses, 2) proinflammatory cytokine IL-12, predominately produced by dendritic cells, is critical in controlling Cryptosporidium infection, and 3) bone marrow-derived CD40-positive cells are required for mice to clear C. parvum infection. Based on these observations, the specific aims are designed to investigate the innate immune response to cryptosporidiosis with a view to determining the mechanisms of Toll-like receptor signaling that lead to such responses, using both mice and human dendritic cells. The specific aims are to: 1) characterize the role of mouse dendritic cells in the innate immune response against C. parvum infection;2) determine the nature of activation of human dendritic cells upon infection with C. parvum or C. hominis;3) identify the Toll-like receptor used by Cryptosporidium and isolate TLR activating ligand(s) expressed by the parasite. The proposed studies will provide the basis for understanding the mechanisms of innate immune recognition and response to parasite invasion necessary for future design of vaccines and other methods of interventions.